INTRODUCTION:

Medicinal preparations take their rightful place in the prevention and treatment for many diseases. In our country, there are more than 800 species of various plants that have medicinal properties. Of these, more than 10% are used in clinical veterinary practice. It should be noted that in many countries up to 50% of drugs are obtained from natural raw materials, mainly from medicinal plants. Studying the experience of traditional medicine and veterinary practice on the use of medicinal plants, of course, helps to increase the number of therapeutic agents in the arsenal of practical medicine and veterinary medicine^1,2.

According to the recommendations of the World Health Organization (WHO), the strategic security of each state is ensured at 20% per share of domestic drugs in the pharmaceutical market. However, on the pharmaceutical market of the Republic of Kazakhstan (RK) with a population of more than 18 million people, domestic manufacturers produce only 10-11% of various herbal medicines that are included in the State Register of the Republic of Kazakhstan³. Compared with other countries and the level recommended by WHO, the domestic industry of Kazakhstan is two times lower than the recommended level. For example, in Spain with a population of 39 million people, the share of herbal preparations is 62%, in France (57 million) -53%, and in Russia (142 million) -35%⁴.

In a comparative aspect, more than 30% of the preparations used in modern medicine are obtained from plant materials. Phytopreparations are used to treat all types of diseases, including socially significant ones such as cardiovascular, oncological, infectious, diseases of the gastrointestinal tract, etc., and, unfortunately, are still not used enough in clinical veterinary medicine^5,6.

Currently, in connection with the deterioration of the environmental situation, it is very important to create new phytopreparations, which, in comparison with synthetic ones, have not only symptomatic, but also more effective pathogenetic effects. The use of antibacterial agents and chemotherapy often leads to the development of various adverse reactions: allergies, getting used to them of pathogenic microflora, suppressing immune reactivity and reducing the efficacy of treatment. Unlike synthetic drugs, phytopreparations have a wider range. They are non-toxic or low-toxic, which makes it possible to use them for a longer time without the risk of any pronounced side effects^7,8.

It should be noted that phytopreparations have other advantages over other methods of healing: environmental safety of plants, allowing long-term and safe use; the evolutionarily established relationship between the active substances of plants and physiologically active substances of the body; polyvalency (versatile orientation) of the action of plants, the possibility of simultaneous treatment by plants of the underlying main and concomitant diseases^9,10.

The plant organism is a very complex creation of nature, both in its chemical composition and in the functions it performs. The diversity of the spectrum of natural chemical compounds contained in plants, and the unique, complex combination of pharmacotherapeutic properties of herbal dosage forms makes it possible to draw up treatment regimens that provide a multilateral, optimal effect for any diseases, taking into account their aetiology and pathogenesis^11,12.

On the basis of the above, it should be stressed that the widespread use of phytopreparations from plant raw materials is particularly important for veterinary medicine, as such preparations possess a wider range of action, fewer side effects and fewer risks of interaction with other preparations. They are much cheaper than synthetic ones and environmentally friendly and can be successfully replaced.

Therefore, the exploration of new phytopreparations from plant raw materials, the study of their pharmacological, immunological and biochemical properties and the introduction into the wide veterinary practice of the world today is of great scientific and practical importance, especially the wealth of the flora of the Republic of Kazakhstan opens up wide prospects in this direction.

In this regard, the study of this issue is undoubtedly a pressing problem at the current stage of the development of veterinary science.

The aim of the work was to develop phytopreparation from local plant raw materials and to study the influence on the indicators of natural resistance of calves of early age.

MATERIALS AND RESEARCH METHODS:

The following medicinal plants were used as a herbal preparation: licorice root (Glyceryhiza glabra), plantain leaves (Plantago glabra), coltsfoot grass (Tussilagofarfara), oregano grass (Origanum vulgare), field horsetail grass (Equisetum arvense)

The chemical composition of these plants is extremely rich in various biological active substances, which have pronounced pharmacotherapeutic properties in various respiratory diseases of humans and animals.

So, licorice root (Fig.1) contains tannins, amino acids, sugar compounds, essential oils, flavonoids, etc. The leaves of plantain contain pectin, flavonoids, tannins, steroid substances, amino acids, essential oil, volatile, vitamins C, other macro- and micro-elements.

Fig.1: Licorice Root (Glyceryhiza glabra)

The leaves of plantain (Plantago glabra) (Fig.2) contain up to 20% of pectin substances, flavonoids, glycosides: aucubin (0.37%) and catalpole, saponins, bitters, tannic, steroid substances, traces of alkaloids; organic acids, amino acids, essential oil, phytoncides, vitamins K, C, macro- and microelements.

Fig. 2: The leaves of plantain (Plantago glabra)

Coltsfoot leaves (Tussilagofarfara) (Fig.3) contain mucous substances, saponins, inulin, tussilagin bitters glucoside, tannins, ascorbic acid, carotenoids (up to 5.18%), gallic, malic and tartaric acids, sitosterol, dextrins, ether oil, flavonoids.

Fig.3: Coltsfoot leaves (Tussilagofarfara)

Oregano grass and flowers (Origanum vulgare) (Fig.4) contain up to 1.2% of ether oil, possessing a bactericidal property. The horsetail grass (Equisetum arvense) contains saponin, organic acids, tannins, flavonoids, amarines, carotene, vitamin C, Ca, K, Fe, Cu, Mg, Zn micro- and macro-elements.

Fig.4: Oregano grass and flowers (Origanum vulgare)

Horsetail grass (Equisetum arvense) (Fig.5) contains saponin, silicic acid, organic acids, tannins, flavonoids, amarines, proteins up to 1.6%, fatty oil up to 3.5%, carotene up to 4.7 mg%, vitamin C 30 to 190 mg%, calcium, potassium, iron, copper, magnesium, zinc.

Fig.5: Horsetail grass (Equisetum arvense)

When collecting medicinal plants, it should be remembered that the value and good quality of plant materials largely depend on their proper drying. The raw materials were dried immediately after collection. Drying medicinal plant materials was carried out with natural (air-solar) heat on the shelves. The average duration of drying was 4 -7 days.

The high quality of plant-based medicinal raw materials depends on many factors and is determined by the correctness and timeliness of its procurement, the content of biologically active substances, and the level of humidity, the absence of mold, impurities and pests. Without quality analyzes, medicinal raw materials are not allowed to be used. Benignness was determined by special research methods, which include the acceptance of raw materials, sampling and their analysis. The authenticity of medicinal plant materials was determined by four analyzes: macroscopic, microscopic, chemical, and luminescent^13,14.

In the course of work, we made collections of medicinal plants (the size of the crushed parts of the plants does not exceed 4-6 mm), before use, we brewed or insisted with water in the ratio 1: 10 (1 part plant, 10 parts water), as well as preparing an alcohol extract (dry extract).

Preclinical and clinical trials were carried out under the conditions of peasant farms of the Almaty region on one-month-old calves of the Alatau cattle breed. The calves were selected taking into account their general condition, age, body weight and were kept in the same conditions of keeping and feeding. Animals were regularly subjected to veterinary control and routine preventive treatments. During the experiments, the farms were safely on infectious and invasive diseases.

The calves were divided into two groups: experimental and control, 5 calves in each group. The calves of the experimental group using a rubber bottle in a mixture with milk were given a phytopreparation in a dose of 70-100ml3, 2 times a day, and the control group was given multivitamin "Tetravit".

Blood for research was taken from the jugular vein in the following sequence: before prescribing the drug (background data), after prescribing it on the 7th, 14th, 21st, 30th days.

Clinical and laboratory blood tests were performed in the clinical diagnostic laboratory of the Sana medical center (Almaty). Quantitative values of immunoglobulins, lysozyme, bactericidal and complementary activity were determined on a Reader StatFax 2100 IFA device, (USA); ELISA automatic analyzer Immulight 1000 (USA), phagocytic activity using microscopes MBL 2000, Karl Zeiss, Prima Star (Germany); M-50, (Austria); total refractometric protein; protein fractions — by agar gel vertical electrophoresis.

The obtained digital data were processed by the constant method of variation statistics with the calculation of arithmetic mean values and their statistical errors (M ± m), the reliability (P) of the compared indicators was determined by the Student criterion. For calculations we used the package of statistical analysis Microsoft Excel.

Change in the protein composition of blood serum is considered to be one of the main indicators showing the general condition of the animal organism.

As a result of our experiments, we obtained data indicating the beneficial effect of the phytopreparation on the protein picture of calf blood serum. These changes affect the content of both the total protein and its individual fractions (Table 1). From the data obtained, it can be seen that before using the herbal preparation, the indicators of the total protein and its individual fractions were approximately at the same level.

Changes began to appear in the subsequent periods of research, especially in the experimental group of calves. In the experimental group, in comparison with the initial data and the control group of calves, a significant increase in the concentration of the total protein and its fractional composition, both in absolute and in relative values, was recorded within the studied periods. Already on the 7th day after giving the phytopreparation, an increase in the level of total protein was observed, where its concentration increased by 2.8% compared to the background value (P<0.05). On 14-21 days, the total protein content began to increase, where the value of the studied parameter rose, respectively, by 5.8 and 8.0% compared with the background data (Р<0.01). It should be noted that the total protein content at the end of the experiment remained at a rather high level in the experimental group of calves.

In the control group of calves, the total protein content remained virtually unchanged at approximately the same level. It was found that the phytopreparation had a pronounced stimulating effect on the dynamics of the content of individual protein fractions. From the research results it was clear that albumin, under the influence of the drug, tended to increase over the entire study period. On the 7th day after prescribing a phytopreparation, the level of albumin compared to the baseline increased by 5.7%; on the 14th day - 13.4%, on the 21st day –19.9%; on the 30th day - 17.7%. (P < 0.001).

Regarding the a- and b- globulin fractions of proteins in the experimental groups, no significant changes were noted. b-globulin fractions of proteins tended to decrease in both experimental and control groups. In contrast, a different pattern was observed in relation to the γ-globulin fraction of proteins. In calves of the experimental group, the level of γ-globulins in the study period constantly tended to increase. So, on the 7th day in comparison with the baseline, the concentration of γ-globulins increased by 3.1%, on the 14th day –10.9%; reaching the highest value on the 21st day, i.e. the degree of increase in this period was 22.4%, and by the end of the experiment - by 16.4% relatively to the background level (P <0.05).

Table 1: The effect of phytopreparation on the dynamics of the protein composition of the blood serum of one-month-old calves, in g/l, (M± m; n = 10)

Indicators	Groups	Background data	Terms of studies, days
Indicators	Groups	Background data	7	14	21	30
Total protein concentration	Ex.	66,12± 1,34^х	68,03 ± 1,12^х	69,94±1,28^х	71,44±1,28^{х х}	70,66±1,39^х
Total protein concentration	C	66,09± 1,28^х	66,14 ± 1,41	66,12±1,09^ххх	66,18±1,14	66,10±1,11
Protein fractions:
Albumin	Ex.	27,18± 0,12^х	28,74 ± 0,09	30,83±0,11^{х х}	32,37±0,12 ^{х х}	31,99±0,11^хх
Albumin	C	27,25± 0,12	27,23 ± 0,12	27,08±0,10	27,18±0,09	27,23±0,09
Globulins	Ex.	38,94± 0,18	39,29 ± 0,14^х	39,11±0,16 ^х	39,07±0,18 ^х	38,67±0,19
Globulins	C	38,84± 0,17^х	38,91 ± 0,17	39,04±0,13 ^{х х}	39,00±0,13^х	38,87±0,13
a-globulin	Ex.	10,98± 0,04	10,57 ± 0,03	9,99±0,03	8,99±0,03	9,19±0,04^хх
a-globulin	C	11,06± 0,05	11,19 ± 0,06^х	11,25±0,04 ^х	11,2±0,05	11,14±0,04
b-globulin	Ex.	10,65± 0,04^х	10,88 ± 0,04^х	9,93±0,04^хх	8,89±0,05	9,33±0,05
b-globulin	C	10,53± 0,04	10,52 ± 0,03	10,52±0,02	10,52±0,03	10,50±0,03^х
γ -globulin	Ex.	17,31± 0,10^х	17,84 ± 0,07^х	19,19±0,09 ^{х х}	21,19±0,10^{х х}	20,15±0,10
γ -globulin	C	17,25± 0,09	17,20 ± 0,08	17,26±0,06	17,2±0,05	17,23±0,06
A/g coef.	Ex.	0,698^х	0,731^{х х}	0,788 ^{х хх}	0,828^{х хх}	0,827
A/g coef.	C	0,702	0,700 ^х	0,694	0,697	0,701

Note: ^хР< 0,05; ^ххР< 0,01; ^х
ххР< 0,001 – relatively to the start of the experiment and in comparison with the control group; Ex.-experiment; C-control; a, b, γ-alpha, beta, gammaglobulins; А/G-albumin / globulincoefficient.

Table 2: The effect of phytopreparation on the indicators of humoral protection of the body of calves (M ± m; n = 10),%

Indicators	Groups	Terms of studies, days
Indicators	Groups	Background data	7	14	21	30
LABS	Ex.	12,32±0,74^х	14,47±0,82^х	18,51±1,12^{х х}	20,69±1,10^{х х}^х	20,54±1,01^х
LABS	C	12,48±0,81^х	12,66±0,96	13,78±1,02^х	13,92±0,87	14,44±0,93^х
BАBS	Ex.	34,12±1,11^х	36,61±1,17^х	39,44±1,08^{х х}	39,56±1,19^х	39,54±1,18
BАBS	C	34,18±1,08	34,52±0,98	35,15±1,22	35,59±1,14	35,51±1,27
CABS	Ex.	10,87±0,49^х	11,88±0,32^х^х	12,12±0,44^х	12,81±0,51^{х х}	12,78±0,48
CABS	C	10,83±0,51	11,09±0.39	11,71±0,56	11,88±0,43	11,88±0,39^х

Note: ^хР<0,05; ^ххР<0,01; ^хххР<0,001 ^хrelatively to the start of the experiment and compared with the control group; LABS, BABS, CABS-lysozyme, bactericidal, complementary activity of blood serum; Ex.-experiment, C-control.

Table 3: The effect of the phytopreparation on the immunoglobulin composition of the blood serum of calves, in mg / ml (М ± m, n = 10)

Indicators	Groups	Background data	Terms of studies, days
Indicators	Groups	Background data	7	14	21	30
IgG	Experiment	21,14±1,16	21,96±1,22^х	23,54±1,39^х	25,81±1,25^ххх	24,79±1,41^х
IgG	Control	21,08±1,08	21,09±1,11	21,19±1,18	21,24±1,20	21,24±1,18
IgM	Experiment	1,79±0,15	1,88±0,18^х	2,22±0,21^хх	2,47±0,31^ххх	2,40±0,27^х
IgM	Control	1,81±0,18	1,83±0,17	1,85±0,14	1,86±0,15	1,85±0,16
IgA	Experiment	0,64±0,11	0,69±0,08^х	0,76±0,09^хх	0,83±0,10^ххх	0,79±0,09^хх
IgA	Control	0,65±0,08	0,65±0,08	0,66±0,07	0,66±0,09	0,67±0,10
Sum of Ig	Experiment	23,57±1,42	24,53±1,48^х	26,52±1,69^х	29,11±1,66^ххх	27,98±1,77^х
Sum of Ig	Control	23,54±1,34	23,57±1,36	23,70±1,39	23,76±1,44	23,76±1,44

Note: ^х Р<0,05; ^хх Р<0,01; ^ххх Р<0,001 ^х relatively to the start of the experiment and in comparison with the control group; IgG, IgM, IgA-immunoglobulinsG, M, A

Fig.6: The effect of phytopreparation on the dynamics of the immunoglobulin composition of calf blood serum, in mg/ml

Table 4: The effect of a phytopreparation on the phagocytic activity of calves’ blood cells and their intensity, (M ± m; n = 10)

Indicators	Groups	Terms of studies, days
Indicators	Groups	Background data	7	14	21	30
PhAL, in %	Experiment	51,3±1,42^х	52,8±1,60	60,5±1,32^{х хх}	60,4±1,24^х	58,2±1,30^{х х}
PhAL, in %	Control	51,7±1,21	52,0±1,44	53,1±1,20^х	53,5±1,18	52,9±1,22
PhI	Experiment	3,39±0,20	3,44±0,24^х	3,76±0,31^{х х}	3,72±0,33	3,62±0,28^х
PhI	Control	3,38±0,22	3,37±0,20	3,41±0,26^х	3,40±0,22	3,35±0,20
PhN	Experiment	2,10±0,18^х	2,14±0,28^х	2,34±0,27^{х хх}	2,33±0,21^х	2,30±0,23^{х х}
PhN	Control	2,11±0,20	2,12±0,20	2,15±0,21	2,14±0,22	2,14±0,18
PhC	Experiment	12220	12640	16040^{х хх}	15145^{х х}	14490
PhC	Control	12210	12280	13900^х	13310	13120

Note: ^х Р<0,05; ^хх Р<0,01; ^ххх Р<0,001 ^х relatively to the start of the experiment and in comparison with the control group; LPhA-leukocytephagocyticactivity; PhI-phagocyticindex; PhN-phagocyticnumber; PhC-phagocyticcapacity

Fig.7: The effect of phytopreparation on the dynamics of the phagocytic activity of leucocytes, in%

The indicators of the control group of animals in all age groups did not undergo special changes and remained at the same level in approximately all periods of the study, with only slight fluctuations.

DISCUSSION OF THE RESEARCH RESULTS:

In order to produce highly productive animals, it is of great importance to produce and grow a full and healthy young man. Since the first days of life, newborns are exposed to various factors of infectious and non-infectious nature, which contribute to the emergence of diseases of the young and reduce the intensity of their growth and development. Diseases occurring in young people usually occur against the background of immunodeficiency, in which there is a reduced response to the action of various agents¹⁵.

There are many ways to increase the protective functions of the body of young animals. One of the promising areas is the development and use of phytopreparations possessing a stimulating and corrective effect on the growth, development of animals and it increases the level of the protective function of young animals¹⁶.

The study of factors of natural resistance will allow differentiating various physiological conditions of the body, as well as the beginning of the development of pathological phenomena. Given all of the above, we set one of the main tasks to study the humoral and cellular factors of the body's natural resistance in young calves.

One of the main indicators characterizing the general condition of the body of young animals is a change in the protein composition of blood serum. Numerous basic studies have proved that proteins are the most important factors of humoral defense, reflect the body’s potential ability to mobilize defenses, and therefore the determination of their activity in the study of natural immunity is of great importance^17,18.

Based on the obtained research results, it should be noted that our phytopreparation has a stimulating effect on the quantitative value of the total protein concentration and protein fractions amount of calves. Therefore, the main effect of the phytopreparation is aimed at biocorrection of metabolic processes in the organs and tissues of the body, which is confirmed by a pronounced increase in the level of total protein and protein fractions concentration¹⁹.

Nowadays, first of all the phenomenon of natural immunity is considered to be the result of the combined activity of humoral and cellular factors ensuring the stability of immunostructural homeostasis of the internal environment under all kinds of the adverse effects. Therefore, the study of factors of immunobiological protection in calves enables to get an objective idea of the potentials of the body in opposing pathogenic agents²⁰.

The studies revealed that the indicators of humoral immunity in calves of the control group were lower than those of the physiological norm and significantly lower than those of the experimental group of calves, which testified an immunodeficiency state.

The obtained data showed that using the phytopreparation significantly activated the indicators of lysozyme, bactericidal and complementary activity of calf blood serum.

Immunoglobulins are known to be one component of humoral immunity. The findings showed that serum immunoglobulin levels in the test group tended to increase compared to the control group of calves at all study periods, apparently due to the intense growth and development of the young calves. The increase in the amount and quality of immunoglobulins indicates the immunocorruption effect of phytopreparation, which in turn has a positive effect on the growth and development of young calves.

An important protective mechanism in the system of immune homeostasis is phagocytosis, which is a reaction of cellular elements in a general biological entity, aimed not only at destroying pathogenic microbes, but also at maintaining cellular balance in the body, both in physiological and pathological conditions. It is one of the factors of both natural congenital and specific acquired immunity. It means that the resistance of young animals, especially from an early age, to various kinds of diseases mostly depends on the level of indicators of cellular immunity of the organism^21,22.

The obtained research results indicate that the use of phytopreparation significantly activates phagocytic cells, which play a decisive role in the system of natural resistance of young animals. Higher rates were found on the 14-21st day after taking the herbal medicine. Studies have shown that experimental calves have lower cellular immunity.

The results of studies on the factors of protective functions of the body of young animals are reliably confirmed by the scientific work of a number of scientists^23,24,25.

CONCLUSION:

Thus, the analysis of the presented data allows us to conclude that calves of early age often have an immunodeficiency condition accompanied by an imbalance of the immune system. Therefore, this circumstance required the used of preparations which actions would be aimed at correcting the violated links of immunity, and our phytopreparation should be believed to meet these requirements.

In order to correct the protective function of the body, we used a phytopreparation from the collection of local raw materials, the action of which, in our opinion, is aimed at biocorrection and bionormalization of metabolic processes of the body of calves. This is confirmed by a significant increase in the level of humoral and cellular factors of immunity protection after the used of phytopreparation.

It should also be stressed that the state of the immune system is the main indicator of the impact of negative environmental factors on the body, and serves as a criterion for the risk of developing many diseases.

Therefore, modern detection of immunodeficiency condition (IDT) and its correction to physiological norm is an important link in the system of increasing natural resistance of the young organism.

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Received on 20.12.2019 Modified on 19.03.2020

Research J. Pharm. and Tech. 2021; 14(2):887-894.

DOI: 10.5958/0974-360X.2021.00158.X

Indicators

Groups

Background data

Terms of studies, days

Terms of studies, days

Control

Control

Control

Control